A variety of protocols have been utilized to induce specific allograft prolongation. These include the use of viable cells, antigen extracts, antibodies and antigen-antibody complexes. In our laboratory, donor bone marrow cells are injected into antilymphocyte (ALS) treated skin grafted recipients and result in graft prolongation over animals treated ith ALS alone. Viable marrow cells mediate their effect in part by stimulating anti-donor blocking factors, decreasing host cell-mediated immunity, and by inducing suppressor T-cells. This effect is specific, since third-party grafts and other antigenic challenges elicit normal host immune responses. The injection of donor marrow for graft prolongation has shown promising results in renal allografts in dogs and in one clinical renal allograft trial. This current approach involves the injection of whole bone marrow i.e., polymorphoneuclear white blood cells, lymphocytes, red blood cells, platelets and other less well defined cells. Preliminary results suggest that not all cell types are necessary for this graft prolongation. Thus, this grant proposal relates to the purification and characterization of a specific cell population from either bone marrow or spleen which is effective in the introduction of graft prolongation. Cell purification methods involve sedimentation of cells at unit gravity in a bovine serum albumin gradient with further cell separation by their nylon wool adherence and electrophoretic properties. Once a homogeneous population is isolated, other methods such as cell receptor analysis, mitogen stimulation index and stem cell assays will be employed for its characterization. Finally, since marrow is injected a week or more after transplantation in this system, it will be tested after storage at minus 180 degrees centigrade. These studies should have a significant impact on organ transplantation since they could (1) show that relatively few cells are necessary for the induction of graft prolongation, (2) lead to other studies related to the mechanism of graft prolongation, (3) establish that frozen isolated cells are able to induce graft prolongation and (4) result in a wide application to human renal transplantation.